High‐throughput quantitative assessment of ABA‐responsive elements at single‐nucleotide resolution

Abstract Abscisic acid (ABA)‐responsive elements (ABREs) are the major cis ‐regulatory elements in ABA‐induced gene expression. However, the impact of sequence variations on ABRE function is not yet well‐understood. Here, we used synthetic STARR‐seq to quantitatively assess the effects of single‐nucleotide substitutions on ABRE activity. Our results revealed that the nucleotide substitutions in both the ACGT‐core and ACGT‐flank regions affected transcriptional strength. Alterations in the ACGT‐core sequence had a more significant impact on ABRE activity than changes in the flanking region. Interestingly, we observed that the ACGT‐flank variants with high activity exhibited a strong sequence preference in the downstream region, whereas the highly active core variants were diverse in sequence patterns. Our studies provide a quantitative map of ABRE activity at single‐nucleotide resolution, which will facilitate the design of ABA‐responsive promoters with desired activities in plants.